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Hemogenic Reprogramming of Human Fibroblasts by Enforced Expression of Transcription Factors
Published on: November 4, 2019
[Expression of coagulation factor IX in human CD34 + hematopoietic stem cells by adeno-associated virus 2]
Yan Chen1, Fang-ping Chen, Jian-qiang Peng
1Department of Hematology, Xiangya Hospital of Central South University, Changsha, China.
Objective:
To investigate the expression of human coagulation factor IX (hFIX) gene in human umbilical cord blood (CB) CD34+ cells which was transfected with recombinant adeno-associated virus 2 (rAAV-2).
Method:
The CD34+ cells were transfected with rAAV-2/hFIX and cultured for 21 days for inducing differentiation into myeloid, erythroid and megakaryocytes, respectively. The expression of hFIX was studied at mRNA, protein and biological activity levels. The cytotoxicity of rAAV-2 to CD34+ cells was evaluated by cell proliferation, cell vitality, CD antigen expressions and CFU yields.
Results:
The hFIX mRNA was expressed in the cultured cells which was verified by RT-PCR and DNA sequencing. An elevated level of hFIX expression and biological activities were detected with a maximum amount of 14.10 ng/10(6) cell x 24 h. During the period of 21 day culture, the cell vitality, cell proliferation, CD antigen expression and CFU yields between the transfected and un-transfected groups had no difference(P > 0.05).
Conclusion:
The human CB CD34+ cells are able to produce functional hFIX after transduction by rAAV-2/hFIX. The cell proliferation and differentiation capacities of the host CD34+ cells were not affected by rAAV-2.
